斑地锦(Euphorbia maculata)是一种常用中药材,其中黄酮类化合物槲皮素是其主要药效成分之一.目前斑地锦槲皮素生物合成基因研究鲜见报道,而实时荧光定量 PCR(RT-qPCR)是基因研究的常用方法之一,但具有合适的内参基因是RT-qPCR相对定量的前提条件.为了获得适合的斑地锦内参基因,本研究基于转录组测序结果,筛选出斑地锦EmSE、EmUBC、EmGDI1、EmLSM12、EmGBP、EmSYP22、EmTRI1、EmRSZ21、EmRPL7以及课题组前期得到的EmUBQ共 10 个基因作为侯选内参基因.通过RT-qPCR检测 10 个候选内参基因在斑地锦营养生长期和生殖生长期的根、茎、叶及果中的表达情况,利用 geNorm、NormFinder 和 BestKeeper 软件分析候选基因的表达稳定性,通过几何平均值法对不同软件所得结果进行整合并明确综合排名,即EmGDI1>EmUBQ>EmSE>EmUBC>EmLSM12=EmGBP>EmTRI1>EmSYP22>EmRSZ21=EmRPL7.以EmGDI1为内参基因,对槲皮素生物合成过程中的一个关键酶基因EmC4H进行表达分析,EmC4H在不同生长期不同组织内的表达差异与转录组测序结果趋势基本一致.本研究开发的内参基因EmGDI1可作为斑地锦不同生长期基因组织特异性表达研究中适合的内参基因.
Erigeron annuus, a member of the Asteraceae family, is widely distributed in tropic, subtropical and temperate regions of China. It is a plant native to North America, which was introduced to China at the end of the nineteenth century as an ornamental plant. E. annuus has great medicinal value. Its whole plant has been used in folk medicine for the treatment of indigestion, enteritis, epidemic hepatitis, lymphadenitis, hematuria, acute inflammation,malaria and obesity. E. annuus could be a phytoremediator for metal-and metalloid-contaminated soils. It can degrade diesel effectively, has strong tolerance and accumulation for uranium, cadmium, zinc, lead, and antimony, and can be distributed in soils with high pollution levels. Hence, In this study, it was to explore the characteristics of chloroplast（cp） genome and its phylogenetic relationship, laying a foundation for the further development of species identification,control and full utilization of E. annuus. The complete cp genome of E. annuus was assembled and sequenced by Illumina Nova Seq 6000 Sequencing platform and SPAdes v3.10. software. Taking the cp genome of E. canadensis（MT806101.1） as a reference, the genome feature and phylogenetic analysis study were investigated. The cp genome of E. annuus was a circular and quadripartite structure with a total length of 153 283 bp. The GC and AT content of the cp genome was 36.77% and 63.23%, respectively. The cp genome of E. annuus had four typical regions, containing a pair of inverted repeats（IRs, 25 019 bp） that were separated by the large single-copy（LSC, 84 833 bp） and small single-copy（SSC, 18 412 bp）. In total, 132 genes were predicted, consisting of 88 protein-coding genes, 36 transfer RNA genes, and8 ribosomal RNA genes. The genes were divided into 4 categories according to the functions, including 45photosynthesis genes, 73 self-replication genes, 6 other genes and 8 genes of unknown function. SSR site analysis revealed that the cp genome sequence contained 200 SSR loci in total, which were spaced disproportionately through the genome with the largest number of SSRs distributed in the LSC region, followed by the SSC and IR regions. LSC, SSC and IR regions had 126, 40, and 34 SSRs, respectively. The number of SSRs in LSC, SSC and IR regions was accounted with 63%, 20% and 17%, respectively. The SSRs were mononucleotides, especially for A/T. The second most abundant motif type was the trinucleotide type, especially TAA, TAT and TTA. Phylogenetic analysis showed that E. annuus was closely related to the plant of E. canadensis. This study revealed the cp genomic information of E. annuus and would provide essential data for further studies on genetics, phylogeny, utilization and prevention of this species. This study revealed the chloroplast genome information and identified the phylogenetic branches of E. annuus, which would provide molecular theoretical support for further research on population genetics, phylogenetic geography, adaptive mechanism and management of E. annuus.
由植原体引起的槟榔黄化病是槟榔的一种毁灭性病害,明确发病槟榔园中植原体遗传多样性及其自然寄主种类,对于全面揭示该病害循环途径及流行规律具有重要意义.本研究对海南不同地区发病槟榔园中表现典型植原体病害症状的植物样品进行调查、采样,利用植原体通用引物进行扩增测序,揭示相关植原体株系的遗传变异规律与系统发育关系.结果表明:从 6 个山黄麻丛枝样品中扩增到植原体 16S rRNA和secA基因片段,不同样品的 2 个基因序列均一致,从 1 个苦楝丛枝样品中扩增到 16S rRNA基因片段.基于 16S rRNA基因序列比对分析表明,山黄麻植原体与已报道山黄麻丛枝植原体序列相似性为 100%,苦楝植原体与已报道苦楝丛枝植原体序列相似性为 100%.系统发育分析表明:山黄麻丛枝植原体与 16SrⅩⅩⅩⅡ组植原体株系聚于一个进化分支,苦楝丛枝植原体与 16SrⅠ组植原体株系聚于一个进化分支;苦楝丛枝植原体与海南已报道槟榔黄化植原体 16S rDNA序列相似性为 100%.及时清理发病槟榔园中苦楝等植原体自然寄主,消除病源,切断植原体的传播途径,对于槟榔黄化病的有效防控至关重要.
天然胶乳稳定剂不仅会影响胶乳的胶体稳定性,对胶膜性能也具有显著影响.本研究选用氢氧化钾(KOH)、氨、十二烷基硫酸钠(SDS)和月桂酸钾 4 种稳定剂添加到浓缩胶乳中,并制备预硫化配合胶乳,研究 4 种稳定剂对浓缩胶乳成膜性能、胶膜硫化特性及物理机械性能的影响.结果表明:4 种稳定剂中对浓缩胶乳胶膜性能影响最大的是KOH,KOH会提高干胶膜的门尼粘度;降低干胶膜收缩率、拉伸强度、定伸应力、断裂伸长率、撕裂强度和硬度等力学性能;同时缩短硫化胶膜的t10、t50 和t90 时间,转矩差值先升后降,定伸应力有所提升,但拉伸强度、断裂伸长率和硬度均会降低,而撕裂强度呈先升后降的趋势.随着氨用量的提高,干胶膜收缩率、拉伸强度、断裂伸长率和撕裂强度均呈先升后降的趋势;硫化胶膜t90 和胶膜收缩率有所升高;拉伸强度、定伸应力同样存在先升后降的趋势.随着SDS用量的提高,浓缩胶乳干胶膜拉伸强度存在先降后升的趋势;硫化胶膜的转矩差值有一定程度的升高;t50 和t90 比较稳定,并在SDS用量超过 0.3%后明显上升,即硫化速度降低;硫化胶膜收缩率快速下降,下降幅度较大.随着月桂酸钾用量的提高,干胶膜门尼粘度和收缩率下降幅度很大;断裂伸长率和撕裂强度有一定提升.硫化胶膜的转矩差值升高幅度最大;胶膜收缩率明显降低,撕裂强度呈先升后降的趋势,其他指标稳定,变化幅度较小.
豇豆枯萎病是由尖孢镰刀菌引起的土传真菌病害,是豇豆最主要的病害之一.利用微生物防治是防控枯萎病的重要手段之一.本研究通过对豇豆-水稻轮作土壤中生防菌的分离、筛选和鉴定,研究其拮抗机制与促生功能,并评估其防治和促生效果,为豇豆枯萎病优良生防菌剂的研制和开发奠定基础.采用平板稀释法分离得到 93 株细菌,再通过平板对峙法筛选出 11 株对豇豆枯萎病病原菌具有抑制作用的拮抗菌株,其中拮抗菌株SD13 抑制率最好(抑制率达到 82.0%);结合形态学与 16S rDNA序列将该菌株鉴定为贝莱斯芽孢杆菌(Bacillus velezensis);通过液体共培养方法发现生防菌SD13 对豇豆枯萎病病原菌的菌丝、孢子均有很强的溶菌作用;采用检测性培养基检测拮抗菌株SD13 抑菌胞外酶(纤维素酶、蛋白酶、几丁质酶、β-1,3-葡聚糖酶)和促生能力相关功能(产嗜铁素、固氮、解无机磷、解有机磷、解钾、产IAA),明确生防菌SD13 可以分泌纤维素酶、蛋白酶、β-1,3-葡聚糖酶 3 种抑菌胞外酶,且具有产嗜铁素、固氮、解无机磷 3 种与促生能力相关的功能;采用平板对峙法测定生防菌SD13 对青瓜枯萎病、辣椒枯萎病等 6 种病原菌的抑制率,结果表明,生防菌 SD13 具有广谱抑菌活性,且抑制率达 66.7%～82.3%;盆栽实验结果表明,在生防菌SD13 的 4 个不同浓度(104、106、108、109 CFU/mL)处理中,106 CFU/mL的防治效果最好达 62.0%,而促生效果仅次于 108 CFU/mL.研究结果表明,生防菌SD13 具有较强的生防与促生效果,并初步明确了其拮抗机制以及与促生能力相关的功能,在防控枯萎病上具有潜在的应用价值.
